Our primary objective is to determine the molecular events that lead to the selective transcription of DNA and subsequent processing and translation of messenger RNA (mRNA). Poxviruses provide a unique and important system for such studies since the enzymes needed for transcription are packaged within the core of infectious virus particles. Purified viral enzymes, including a guanylyltransferase and two methyltransferases have been used to reconstruct the sequence of reactions leading to the methylated cap structure of vaccinia virus mRNA, to investigate the roles of the individual modifications on ribosome binding and protein synthesis, and for RNA sequencing. Similar and additional enzymes have been isolated from uninfected Hela cells to determine the steps involved in the processing of eukaryotic mRNA. Evidence for temporal control of vaccinia virus gene expression at the transcriptional level has been obtained by DNA:RNA hybridization studies and by translation of viral mRNA in a cell-free protein synthesizing system. A transcriptional map of the vaccinia virus genome is being prepared using DNA fragments produced by restriction endonuclease digestion. Electron microscopic analysis of the vaccinia virus genome led to the visualization of a 10,500 nucleotide inverted terminal repetition. Preliminary studies suggest that a portion of the repeated sequence is transcribed.